Dear Bomba
I am bit confused , Are you running bowtie ---> tophat ----> Cufflink
For bowtie run you are using reference genome annotation in gff3 format
I am curremntly totally unable to figure it out correctly, how i should analyse my RNA
seq data
Best
Ateeq
________________________________
From: Noa Sher <noa.sher(a)gmail.com>
To: Bomba Dam <bomba.dam(a)visva-bharati.ac.in>
Cc: galaxy-user(a)lists.bx.psu.edu
Sent: Wednesday, February 22, 2012 7:28 AM
Subject: Re: [galaxy-user] Cufflinks related problem
Hi Bomba
I cant know for sure without seeing your files but I originally had the same problem and
it ended up being because the way the genome was named in the fasta genome file was not
the same as the way it was named in column 1 of my gtf file. I would check that first.
Also- with bacteria you dont want to run cufflinks with default parameters- use the galaxy
browser to check how your data looks after tophat and you will most likely see very
strange gene-spanning introns. Change the max intron size to 1000-1500 and the min
distnace to 101-5nt and you should get results that make much more sense.
Good luck
Noa
On 22/02/2012 00:12, Bomba Dam wrote:
Dear Noa,
Can you please tell me the parameters that I should keep while
mapping
bacterial transcripts using cuffkins. I have kept the
default parameters as in Cufflinks and used my custom genome
annotation in gff3 format. The cufflinks seems to work Ok but all
the FPKM values in these files are zero. As suggested by other
users I have checked the correctness of my GFF3 files. The
corresponding fasta file was used for mapping the transcripts
using Bowtie. Are there any special trick for mapping bacterial
transcriptome.
regards,
Bomba
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